Deputy Australian Greens leader Larissa Waters has resigned from the Senate after realising she has been ineligible to sit in Federal Parliament since 2011.
Ms Waters was born in Canada and did not renounce her dual citizenship when running for office.
The country's constitution disqualifies potential candidates from election if they hold citizenship in other countries.
The Queensland senator's resignation comes less than a week after Greens senator Scott Ludlam quit after realising he held dual New Zealand citizenship.
The resignations mean the Greens have lost two of their strongest performers in Australian Parliament in less than a week.
Ms Waters' resignation is effective immediately.
The former Queensland senator was tearful when she apologised to her constituents for her oversight.
In a statement, Ms Waters said she had not visited Canada since leaving as a baby and believed she had naturalised to Australia.
She sought legal advice after Mr Ludlam's resignation and said she was shocked and saddened by the result.
"I was devastated to learn that because of 70-year-old Canadian laws I had been a dual citizen from birth, and that Canadian law changed a week after I was born and required me to have actively renounced Canadian citizenship," she said.
"It is with a heavy heart that I am forced to resign as senator for Queensland and co-deputy leader of the Australian Greens, effective today.
"I apologise wholeheartedly to all those who have supported me and helped me to become a representative for the wonderful people of Queensland over the last six years."
Ms Waters was seen by some in the party, including former leader Bob Brown, as a future leader of the party.
She was made co-deputy leader of the party in May 2015 and has been a strong opponent of the proposed Adani mine in the Galilee Basin.
She made international headlines this year when she became the first woman to breastfeed her daughter, Alia, on the floor of the country's Parliament.
Her resignation means the party is facing two senate recounts in Queensland and West Australia.
Former leader of the Australian Democrats Andrew Bartlett is in line to re-enter the Senate as he would likely win a countback after Ms Waters resignation.